Apoptosis is programmed cell death, which is characterized by condensation of the nucleus and cytoplasm, specific protein degradation, and DNA fragmentation. Extensive studies have documented that apoptosis is the primary killing mechanism induced by cancer therapies such as chemotherapy, radiotherapy or the cell death factor TNFalpha. Oral cancer is a widely distributed cancer which is typified by squamous cell carcinoma (SCC). Importantly, SCC is very resistant to cancer therapy and TNFalpha, but the genetic and molecular mechanisms underlying this resistance are complex and poorly understood. Our preliminary studies from in vitro and in vivo presented in this application indicate that NF-kB is a pro-survival transcription factor for oral squamous cell carcinoma and inhibition of NF-kB renders SCC sensitive to TNFalpha-mediated apoptosis. Using TNFalpha as death inducer, the cell death machinery of SCC will be systematically dissected, and the molecular mechanism of NF-kB anti-apoptosis will be explored. The specific aims proposed in this application are to: 1) Determine the primary mechanisms of SCC resistance to TNFalpha-mediated apoptosis by examining how NF-kB inducible anti-apoptotic genes inhibit caspase-8 activation; 2) Explore whether activation of NF-kB directly regulates TNFalpha-mediated apoptosis at the level of mitochondria in SCC, focusing on the role of NF-kB regulated A1 and manganese superoxide dismutase (MSN) proteins in inhibition of the release of cytochrome c; 3) Determine the expression of NF-kB inducible anti-apoptotic genes in SCC in vivo by immunohistochemistry, and thereby explore whether the aberrant expression of specific anti-apoptotic genes is related to the development of SCC; and 4) Identify potential regulators of apoptosis of SCC by functional cloning, and explore the potential role of these regulators in the development of SCC. These studies will elucidate the intrinsic genetic and molecular mechanism of cell resistance to TNFalpha-mediated apoptosis. Importantly, these studies will help to develop the biological marker for the diagnosis of oral cancer and provide a molecular basis for improving efficacy of cancer therapy by manipulating the apoptotic program of oral cancer cells.